PROJECT SUMMARY Transposable elements (TEs) are genomic parasites that can negatively impact host viability and fertility. Despite the detrimental effects of TEs, they constitute appreciable proportions of virtually all eukaryotic genomes surveyed. Eukaryotic genomes can be cytologically divided into euchromatin and heterochromatin, with the latter being tightly packaged DNA generally associated with suppressed expression. While the heterochromatin of most species is infested with degenerated TEs, the proportions of host euchromatic genomes that are occupied by potentially active TEs are remarkably different between species (e.g., human, 45%; Drosophila melanogaster, 5.4%). This proposal aims to understand the molecular and evolutionary mechanisms contributing to the widely observed, but poorly understood, between-species differences in TE content, which will be critical for the understanding of eukaryotic genome evolution and the genetics of inherited diseases and cancers caused by TEs. Natural selection can remove deleterious TE insertions from populations, and variation in the strength of selection is expected to result in differences in TE content between host genomes. Most previous work on the deleterious effects of TEs has centered on the consequences of TE-induced physical disruption of DNA. Instead, this proposal focuses on the largely unexplored epigenetic effects of TEs. Constitutive heterochromatin has the potential to influence the epigenetic states of adjacent genes through cis-spreading of repressive epigenetic marks as well as spatial interactions with other heterochromatic regions. In euchromatin, active TEs can be silenced through host-directed enrichment of repressive epigenetic marks, which were also observed to spread in cis into adjacent genes and thereby epigenetically impair gene function. By using Drosophila as a model, this proposal will use a combination of Hi-C analysis and cell biological experiments to investigate if euchromatic TEs epigenetically impact adjacent genes through spatial interactions with constitutive heterochromatin, which will identify previously unknown epigenetic effects of TEs (Aim 1). Host genotypes and environmental conditions are known to modulate the spreading of silencing marks from heterochromatin to adjacent sequences. The influence of these host components on the epigenetic effects of TEs in euchromatin will be tested (Aim 2). Finally, this proposal will use comparative functional genomics and phylogenetic analysis to test the hypothesis that between-species variation of host components that modulate TE's epigenetic effects contributes to different epigenetic effects of TEs, and ultimately divergent TE content across the Drosophila phylogeny.